α4 integrins are prominent on mononuclear leukocytes. They mediate cell migration, which is dependent on binding of the intracellular adaptor protein paxillin to the cytoplasmic tail of α4.
α4 integrin antagonists are effective in inhibiting a wide variety of experimental models of inflammatory diseases(1-4) and autoimmunity because they inhibit the recruitment of lymphocytes and monocytes to sites of inflammation. Furthermore, anti-α4 antibodies are of proven therapeutic effectiveness in human autoimmune diseases, such as Multiple Sclerosis(5). Anti-α4 antibodies, such as natalizumab, and small molecule and peptidomimetic α4 antagonists inhibit the integrin's interactions with ligands such as VCAM-1. At saturation, this form of inhibition causes complete loss of α4 integrin function. Consequently, use of these agents recapitulates the null phenotype implying the potential for mechanism-based toxicities such as defects in placentation, heart development, and hematopoiesis(6). Furthermore, the blockade of T cell entry into the central nervous system accounts for the occurrence of Progressive Multifocal Leukoencephalopathy in humans treated with anti-α4 integrin antibodies(7).
Integrin functions depend on their capacity to generate and respond to cellular signals. Blockade of integrin signaling can leave ligand binding function intact(8-10). Consequently, only partial inhibition of function may occur, even with full blockade of the target, potentially providing a more favorable therapeutic window. A quest for interactions important in α4 integrin signaling uncovered a tight binding interaction of the α4 cytoplasmic domain with paxillin, a signaling adaptor (11). Blocking the interaction by mutations of α4 (e.g. α4(Y991A)) that selectively block paxillin binding, reduces cell migration(10;11).
Furthermore, inhibition of paxillin binding to α4 by a fragment of paxillin(12) or a small molecule antagonist(13) also impairs migration, suggesting that such agents could be used as therapeutic inhibitors of α4 integrin function. Notably, blocking the interaction of α4 with paxillin does not disrupt α4-mediated static adhesion(8;11), suggesting that this form of antagonism might not interfere with functions such as anchorage of hematopoietic progenitors in the bone marrow. Present inventors have previously confirmed this idea by generating mice homozygous for an α4 mutation (α4(Y991A)) that selectively(11) blocks paxillin binding. Similarly, inventors have previously reported that unlike α4 null mice(6;14), these α4(Y991A) mice are viable and fertile; however, they manifest a profound deficit in the recruitment of mononuclear leukocytes to an inflammatory site with no defect in neutrophil recruitment(29). Furthermore, α4 integrins are essential for definitive hematopoiesis(6;15;16); however, the α4(Y991A) mice exhibited normal hemograms, normal abundance of hematopoietic precursors and unimpaired homing of hematopoietic progenitor cells to the bone marrow, a surrogate marker of stem cell migration. Thus inventors have previously established the principle that blockade of α4 integrin signaling can impair mononuclear leukocyte recruitment to an inflammatory site, while averting the adverse effects of α4 integrin loss on development and hematopoiesis (29).
While existing antagonists of the α4 integrin are useful as therapeutic agents for autoimmune diseases, they are also associated with a number of serious toxic side effects. Specifically, known antagonists of the α4 integrin can affect development and hematopoiesis, and are also associated with progressive multifocal leukoencephalopathy.
The present invention describes novel compounds that are able to inhibit the interaction of paxillin or its paralogues with α4 integrin or its binding partners that regulate signaling events downstream of the paxillin-α4 interaction and methods for therapeutic use of such compounds. The compounds of the present invention are able to inhibit the interaction of paxillin or its paralogues with α4 integrin or its binding partners without resulting in hematopoietic or developmental toxicities.